Ultrasensitive quantification of PD-L1+ extracellular vesicles in melanoma patient plasma using a parallelized high throughput droplet digital assay.

The expression of programmed death-ligand 1 (PD-L1) on extracellular vesicles (EVs) is an emerging biomarker for cancer, and has gained particular interest for its role mediating immunotherapy. However, precise quantification of PD-L1+ EVs in clinical samples remains challenging due to their sparse concentration and the enormity of the number of background EVs in human plasma, limiting applicability of conventional approaches. In this study, we develop a high-throughput droplet-based extracellular vesicle analysis (DEVA) assay for ultrasensitive quantification of EVs in plasma that are dual positive for both PD-L1 and tetraspanin (CD81) known to be expressed on EVs.

Parallelized immunomagnetic nanopore sorting: modeling, scaling, and optimization of surface marker specific isolation of extracellular vesicles from complex media.

The isolation of specific subpopulations of extracellular vesicles (EVs) based on their expression of surface markers poses a significant challenge due to their nanoscale size (< 800 nm), their heterogeneous surface marker expression, and the vast number of background EVs present in clinical specimens (10 -10 EVs/mL in blood). Highly parallelized nanomagnetic sorting using track etched magnetic nanopore (TENPO) chips has achieved precise immunospecific sorting with high throughput and resilience to clogging. However, there has not yet been a systematic study of the design parameters that control the trade-offs in throughput, target EV recovery, and specificity in this approach.

Extracellular Vesicle-Based Multianalyte Liquid Biopsy as a Diagnostic for Cancer.

Liquid biopsy is the analysis of materials shed by tumors into circulation, such as circulating tumor cells, nucleic acids, and extracellular vesicles (EVs), for the diagnosis and management of cancer. These assays have rapidly evolved with recent FDA approvals of single biomarkers in patients with advanced metastatic disease. However, they have lacked sensitivity or specificity as a diagnostic in early-stage cancer, primarily due to low concentrations in circulating plasma.